脑出血炎症反应上调铁调素的表达影响脑铁清除及机制

Reducing the deposited brain iron is one of important measures in treatment of intracerebral hemorrhage (ICH). Currently, as the mechanism of brain iron metabolism and its regulating and controlling factors after ICH are not clear, the effect of the treatment of reducing iron simply by physical method has larger difference. So exploring the mechanism of brain iron metabolism following ICH and giving the targeted intervention have important roles in the ICH therapy. Given that hepcidin plays an important role in iron metabolism, our pre-experiments has also found that the astrocyte expression of hepcidin was significantly increased after ICH, while both of the hepcidin expression levels and the brain iron contents around the hematoma brain tissue were reduced in the TLR4 knockout mice ICH model. Combined with the studies that have confirmed the interactions of the hepcidin and the ferroportin (FPN) expressed on the vascular endothelial cell membrane plays an important role in iron metabolism, thus we put forward the following hypothesis: TLR4 mediated the inflammatory response upregulated the astrocytes expression of hepcidin after ICH, then combined with FPN expressed on the endothelial cell membranes slowing the deposited brain iron's expelling. Therefore based on this, we proposed the further study aims: (1) to explore the role of hepcidin in the brain iron metabolism after ICH; (2) to study the mechanism of the upregulated hepcidin following ICH; (3) to investigate whether TLR4 inhibitors could improve the effect of ICH treatment via inhibiting the hepcidin expression to reduce the deposited brain iron. The study can further elucidate the mechanism of brain iron metabolism after ICH, and is expected to provide new methods for the treatment of ICH.

降低脑铁沉积是脑出血（ICH）治疗的重要手段，由于目前ICH后脑铁代谢机制及其调控因素不清楚，单纯通过物理方法降铁治疗的效果存在较大差异。因此探索ICH后脑铁代谢机制并进行针对性干预有重要意义。鉴于铁调素在铁代谢中起着重要作用，我们预实验发现ICH后星形胶质细胞铁调素表达显著上调，而且还发现TLR4敲除鼠ICH后铁调素表达及脑铁含量均降低。结合已有研究证实铁调素与血管内皮细胞膜上膜铁转运蛋白（FPN）相互作用在铁代谢中具有重要作用，据此我们提出如下假说：ICH后TLR4介导的炎症反应上调星形胶质细胞铁调素表达，进而与内皮细胞膜上FPN结合，导致脑铁清除受阻。为此本项目拟进行：（1）明确铁调素在ICH后脑铁代谢中的作用；（2）探讨ICH后铁调素表达上调的机制；（3）探索通过抑制铁调素表达、降低脑铁含量对ICH治疗的效果。该研究可进一步阐明ICH后脑铁代谢的机制，可望为ICH的治疗提供新方法。

脑出血后脑铁沉积是导致脑损伤加重的重要原因之一，因此促进沉积脑铁清除是减轻脑出血脑损伤的重要手段之一。尽管已有研究报告通过应用铁螯合剂来降低脑铁沉积，进而减轻脑损伤，但其在动物模型及远期的疗效不确切。这可能与脑出血后脑铁代谢机制不清楚有关，因此阐明脑出血后脑铁代谢机制有望为促进沉积脑铁清除治疗脑出血提供新靶点。鉴于铁调素是贴调节的重要激素之一，在铁代谢中发挥重要作用。为此，在本项目的资助下，我们首先利用脑出血患者血清标本，采用铁调素ELISA试剂盒检测脑出血患者血清铁调素水平，发现预后差的脑出血患者其血清铁调素水平显著高于预后好的患者；进一步利用多元线性回归分析后发现血清铁调素含量越高，脑出血患者临床结局预后越差，提示铁调素在脑出血脑铁代谢中具有重要的作用。在此基础上，我们利用小鼠并联共生+脑出血模型，发现小鼠脑出血后脑组织铁调素表达显著上调，在第3天达到高峰，并且主要由星形胶质细胞表达，同时血清铁调素含量亦显著增高。进一步利用铁调素敲除鼠，发现与野生鼠相比，脑出血后脑铁含量显著降低、脑组织氧化损伤及认知功能障碍等均得到显著改善。然而，向野生鼠静脉注射铁调素加重上述指标。离体实验表明，星形胶质细胞表达的铁调素通过与脑微血管内皮细胞（BMVECs）膜上的膜铁转运蛋白（FPN1）结合导致BMVECs胞内铁排出减少，这种现象可以被铁调素拮抗剂fursultiamine逆转。利用并联共生脑出血模型，发现外周升高的血清铁调素亦可显著阻碍脑出血后沉积脑铁清除），阐明了脑出血远期认知功能障碍的发生机制。我们进一步的研究发现脑出血后激活的TLR4/MyD88信号产生IL-6，通过JAK/STAT3途径促进铁调素表达。检测发现TLR4-/-, MyD88-/-小鼠脑出血后第7、14及28天脑铁含量较TRIF-/-及野生小鼠显著降低。并且，连续5天腹腔注射TLR4拮抗剂TAK242亦显著降低了野生鼠脑出血后第14及28天脑铁含量，并且改善脑出血小鼠28天时的认知功能障碍，提示脑出血后炎症反应在调控脑铁代谢中具有重要的作用。